Tuyen Do Gia, Kitamura Kenichiro, Adachi Masataka, Miyoshi Taku, Wakida Naoki, Nagano Junko, Nonoguchi Hiroshi, Tomita Kimio
Department of Nephrology, Kumamoto University Graduate School of Medical Sciences, Kumamoto, Japan.
Kidney Int. 2005 Jan;67(1):193-200. doi: 10.1111/j.1523-1755.2005.00070.x.
Prostasin has been shown to be involved in the regulation of sodium handling in the kidney. TGF-beta1 has been demonstrated to suppress alphaENaC expression and sodium uptake. Therefore, we hypothesized that TGF-beta1 may regulate prostasin expression to modulate sodium reabsorption in the kidney.
To determine if TGF-beta1 has an effect on prostasin expression, we isolated 2.9 kb of the rat prostasin promoter, and measured its transcriptional activity with a luciferase assay in mouse cortical collecting duct cell line (M-1). The effect of TGF-beta1 on the mRNA and protein abundance of prostasin, and amiloride-sensitive (22)Na uptake was determined.
Treatment of M-1 cells with 20 ng/mL of TGF-beta1 for 24 hours significantly decreased the promoter activity by 50 +/- 1%, and the inhibitory effect was dose dependent over the range of 0.1 to 20 ng/mL. We identified a 50 bp region (-410 to -360) containing c-Rel-like sequence in prostasin promoter that is responsible for the TGF-beta1-mediated inhibition, and found that TGF-beta1 increases IkappaBalpha expression in M-1 cells. TGF-beta1 reduced endogenous prostasin mRNA and protein expression in M-1 cells by 50 +/- 12% and 44 +/- 12%, respectively, and the amiloride-sensitive (22)Na uptake by 35.9 +/- 4.8%.
Our findings indicate the possibility that TGF-beta1 transcriptionally inhibits prostasin expression by the induction of IkappaBalpha and the subsequent inhibition of NF-kappaB/Rel activity in M-1 cells, and also suggest the possibility that TGF-beta1 might inhibit sodium reabsorption through a reduction in prostasin expression and subsequent inhibition of ENaC activity.
已表明前列腺素酶参与肾脏中钠处理的调节。转化生长因子-β1（TGF-β1）已被证明可抑制α-上皮钠通道（αENaC）表达和钠摄取。因此，我们推测TGF-β1可能调节前列腺素酶表达以调节肾脏中的钠重吸收。
为了确定TGF-β1是否对前列腺素酶表达有影响，我们分离了大鼠前列腺素酶启动子的2.9 kb片段，并在小鼠皮质集合管细胞系（M-1）中用荧光素酶测定法测量其转录活性。确定了TGF-β1对前列腺素酶的mRNA和蛋白质丰度以及氨氯地平敏感的（22）钠摄取的影响。
用20 ng/mL的TGF-β1处理M-1细胞24小时，可使启动子活性显著降低50±1%，且在0.1至20 ng/mL范围内，抑制作用呈剂量依赖性。我们在前列腺素酶启动子中鉴定出一个包含c-Rel样序列的50 bp区域（-410至-360），该区域负责TGF-β1介导的抑制作用，并发现TGF-β1可增加M-1细胞中IκBα的表达。TGF-β1使M-1细胞中内源性前列腺素酶mRNA和蛋白质表达分别降低50±12%和44±12%，并使氨氯地平敏感的（22）钠摄取降低35.9±4.8%。
我们的研究结果表明，TGF-β1可能通过诱导IκBα并随后抑制M-1细胞中的NF-κB/Rel活性，在转录水平上抑制前列腺素酶表达，还提示TGF-β1可能通过降低前列腺素酶表达并随后抑制ENaC活性来抑制钠重吸收。
